1. Technical Field
The present invention relates to a method for producing L-amino acid, such as L-histidine. More specifically, the present invention relates to a novel feedback-resistant enzyme involved in the biosynthesis of purines and L-histidine. More specifically, the present invention concerns a new feedback-resistant mutant phosphoribosylpyrophosphate synthetase (PRPP synthetase) from E. coli. The invention also relates to a method for producing L-histidine by fermentation using bacterial strains containing the novel feedback-resistant enzyme.
2. Background Art
Conventionally, L-amino acids are industrially produced by fermentation methods utilizing strains of microorganisms obtained from natural sources or mutants thereof, which are modified to enhance production yields of L-amino acids.
Many techniques to enhance production yields of L-amino acids have been reported, including transformation of microorganisms with recombinant DNA (see, for example, U.S. Pat. No. 4,278,765). Other techniques include increasing the activities of enzymes involved in amino acid biosynthesis and/or desensitizing the target enzymes of the feedback inhibition by the resulting L-amino acid (see, for example, Japanese Laid-open application No. 56-18596 (1981), WO 95/16042 or U.S. Pat. Nos. 5,661,012 and 6,040,160).
5-Phosphoribosyl-α-1-pyrophosphate (hereinafter, “PRPP”) and adenosine-5′-triphosphate (hereinafter, “ATP”) are the initial substrates in histidine biosynthesis. PRPP can sometimes induce the histidine biosynthesis to follow divergent pathways, resulting in the biosynthesis of pyrimidine nucleotides, purine nucleotides, pyridine nucleotides, and tryptophan (Escherichia coli and Salmonella, Second Edition, Editor in Chief: F. C. Neidhardt, ASM Press, Washington D.C., 1996).
Many nucleotides competitively inhibit the activity of PRPP synthetase with ATP. However, the only potent nucleotide inhibitor is adenosine-5′-diphosphate (ADP); it competes with ATP and is an allosteric inhibitor that binds to a site other than the active site (Hove-Jensen, B. et al, J. Biol. Chem. 261:6765-6771 (1986)).
Mutants with altered PRPP synthetase have been obtained in both E. coli and S. typhimurium. One of the E. coli mutants produces a PRPP synthetase with a 27-fold increase in the Km value for ATP, and the enzyme is no longer inhibited by AMP. This mutation results from substitution of aspartic acid 128 by alanine (prsDA mutation). One S. typhimurium prs mutant is temperature-sensitive and has only 20% of the wild-type PRPP synthetase activity. This mutant enzyme had elevated Km values for ATP and ribose 5-phosphate and reduced sensitivity to inhibition by ADP. The mutation is the result of the replacement of arginine 78 by cysteine (Escherichia coli and Salmonella, Second Edition, Editor in Chief: F. C. Neidhardt, ASM Press, Washington D.C., 1996).
It is well known that superactivity of human PRPP synthetase and resistance to purine nucleotide are associated with neurodevelopmental abnormalities in addition to hyperuricemia and gout (Becker M. A. et al, Arthritis Rheum, 18:6 Suppl: 687-94 (1975); Zoref E. et al, J. Clin. Invest., 56(5): 1093-9 (1975)). Uric acid overproduction in individuals with superactivity of PRPP synthetase results from increased production of PRPP and consequent acceleration of purine nucleotide synthesis de novo. It was shown that superactivity of PRPP synthetase is a result of an A to G mutation at nucleotide 341, which results in an asparagine to serine substitution at amino acid residue 113 of the mature enzyme. This mutant PRPP synthetase is resistant to purine nucleotides that inhibit the normal enzyme by a mechanism that is noncompetitive with respect to ATP (Roessler, B. J. et al. J. Biol. Chem., v. 268, No 35, 26476-26481 (1993); Becker, M. A. et al, J. Clin. Invest., 96(5): 2133-41 (1995)).
A process for producing purine nucleosides via fermentation of a microorganism belonging to the genus Escherichia and having purine nucleoside-producing ability, and containing a prsDA mutation is disclosed (European patent application EP1004663A1). However, there are no reports describing mutant bacterial PRPP synthetase which is feedback-resistant to purine nucleotides, or the use of such a mutant PRPP synthetase for improving L-histidine production using L-histidine-producing strains.